Structural Biology outside the box—inside the cell

•Biophysical methods for studying proteins in situ reached unprecedented levels of maturity.•Direct investigations of cellular protein structures and functions are now possible.•Future efforts will capitalize on complementary aspects of these methods and their joint application. Recent developments in cellular cryo-electron tomography, in-cell single-molecule Förster resonance energy transfer-spectroscopy, nuclear magnetic resonance-spectroscopy and electron paramagnetic resonance-spectroscopy delivered unprecedented insights into the inner workings of cells. Here, we review complementary aspects of these methods and provide an outlook toward joint applications in the future.